title: Indirect activation of the SV23 and SV24 splice variants of human constitutive androstane receptor: analysis with 3-hydroxyflavone and its analogues.
abstract: OBJECTIVE
  Naturally occurring splice variants of human CAR (hCAR), including hCAR-SV23 (insertion of amino acids SPTV) and hCAR-SV24 (APYLT), have been shown to be expressed in liver. However, little is known regarding how hCAR-SV23 and hCAR-SV24 are activated. Therefore, we investigated the mode of activation of these hCAR splice variants.
  
  
  METHODS
  Cell-based reporter gene assays, including ligand-binding domain transactivation assays and coactivator recruitment assays, were conducted on cultured HepG2 cells transfected with various constructs and treated with 3-hydroxyflavone or a hydroxylated (galangin, datiscetin, kaempferol, morin, quercetin or myricetin) or methylated (isorhamnetin, tamarixetin, or syringetin) analogue.
  
  
  RESULTS
  Among the flavonols investigated, only 3-hydroxyflavone increased hCAR-SV23 and hCAR-SV24 activities. 3-Hydroxyflavone did not transactivate the ligand-binding domain of these isoforms or recruit steroid receptor coactivators (SRC-1, SRC-2, or SRC-3). By comparison, 3-hydroxyflavone, galangin, datiscetin, kaempferol, quercetin, isorhamnetin and tamarixetin activated hCAR-WT, whereas none of the flavonols activated hCAR-SV25 (both SPTV and APYLT insertions). The flavonols 3-Hydroxyflavone, galangin, quercetin and tamarixetin transactivated the ligand-binding domain of hCAR-WT, but only 3-hydroxyflavone recruited SRC-1, SRC-2 and SRC-3 to the receptor.
  
  
  CONCLUSIONS
  hCAR-SV23 and hCAR-SV24 can be activated by a mechanism that does not involve the ligand-binding domain of the receptor or recruitment of SRC-1, SRC-2, or SRC-3. 3-Hydroxyflavone and its structural analogues activated hCAR in an isoform-selective and chemical-specific manner. Overall, our study provides insight into a novel mode of ligand activation of hCAR-SV23 and hCAR-SV24.
authors.affiliation: Faculty of Pharmaceutical Sciences, The University of British Columbia, Vancouver, BC, Canada.
authors.forename: Aik Jiang
authors.initials: AJ
authors.lastname: Lau
authors.forename: Thomas K H
authors.initials: TK
authors.lastname: Chang
mesh_terms: D000090702:Constitutive Androstane Receptor; D005419:Flavonoids / Q000737:chemistry / Q000494:pharmacology*; D017930:Genes, Reporter; D056945:Hep G2 Cells; D006801:Humans; D008024:Ligands; D008099:Liver / Q000378:metabolism*; D056920:Nuclear Receptor Coactivator 1 / Q000378:metabolism; D051958:Nuclear Receptor Coactivator 2 / Q000378:metabolism; D056921:Nuclear Receptor Coactivator 3 / Q000378:metabolism; D020033:Protein Isoforms; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D014162:Transfection
chemical_list: D000090702:Constitutive Androstane Receptor; D005419:Flavonoids; D008024:Ligands; D051958:Nuclear Receptor Coactivator 2; D020033:Protein Isoforms; D018160:Receptors, Cytoplasmic and Nuclear; D056920:Nuclear Receptor Coactivator 1; D056921:Nuclear Receptor Coactivator 3; C041477:3-hydroxyflavone
keywords: 3-hydroxyflavone; constitutive androstane receptor; flavonols; splice variants; steroid receptor coactivators